The cytochalasins are a group of cytotoxic fungal metabolites which exhibit a complex array of biological activities. These molecules have been used extensively as probes of cell biological processes. The specific objectives of this investigation are: (1) the design and synthesis of simplified cytochalasin analogs, into which 13C-labels, fluorophores, affinity labeling functionality and/or modified collections of functional groups have been incorporated; (2) the determination of specific structure-activity correlations of cytochalasins; (3) determination of specific molecular aspects of the mechanism of actin polymerization; (4) identification and characterization of cellular cytochalasin receptors. We will develop versatile synthetic strategies based primarily upon established methodology from total synthetic studies of cytochalasins, but which branch from well-developed key intermediates. Biological activity will be assessed (in collaborative efforts) by two techniques: (1) a cell-free gelation assay; and (2) a laser fluorescence photobleaching recovery assay. NMR and FPR studies of simplified cytochalasins will determine the mechanisms of actin polymerization and specific molecular interaction of cytochalasins with actin. Affinity labeling studies will identify cytochalasin binding sites which mediate cellular responses to cytochalasins. We also anticipate that simplified cytochalasin analogs with narrow, more well-defined spectra of biological activity will find extensive applications as finetuned probes of membrane and cytoskeleton-associated biological processes.